Metastasis is the process by which blood-borne cancer cells establish new tumor colonies in select, secondary organs. Using lung-metastatic human and mouse breast cancer cells, we have shown that these tumor cells form high-affinity bonds with lung endothelium via the endothelial cell adhesion molecule CLCA2 or (mClca2) and its tumor ligand the beta4 intregin. Beyond causing vascular arrest, this adhesion initiates the steps that are necessary for cancer cells to penetrate the endothelial cell lining and to form secondary tumor colonies outside the vascular compartment. In this proposal, we will explore the effects of the CLCA2/beta4 adhesion on the bound cell pair and assess the contributions of the two cells to extravasation. The endothelial cell contribution will be evaluated by investigating the role of structural peculiarities of CLCA2, including the beta4 -binding domain, an N-terminal extracellular Zn-finger-like domain, and a proteolytic processing site that is responsible for cleavage of a primary CLCA2 translation product into two mature, plasma membrane-associated proteins, in proper CLCA2 expression and function as well as downstream signaling in preparation of tumor cell extravasation. The tumor cell contribution will be assessed by exploring the central role of CLCA2 /beta4-mediated activation of focal adhesion kinase in tumor cell migration and proliferation, both essential for successful extravasation. However, of particular interest will be the dualistic function of CLCA2 as an adhesion molecule and a Cl channel. The question will be asked whether a suicide pathway is activated in endothelial cells by adherent tumor cells that involves stimulation of endothelial Cl conductance, resulting in "holes" in the endothelial cell monolayer that are used by tumor cells to traverse the endothelial lining. These experiments will be complemented by studies of beta4 expression during breast cancer progression in a human xenograft model, the contribution of CLCA2 to tissue selection of metastasis and metastasis development, and the generation of a mouse model with conditional expression of mClca2 in endothelia of adult mice to study susceptibility to metastasis. Understanding the molecular principles that govern extravasation will ultimately allow us to develop strategies in anti-metastatic therapies.